GSM6812992: sPAP_m1; Mycobacterium tuberculosis H37Rv; ncRNA-Seq
RNA was isolated using TRIzol LS reagent (Invitrogen). Cells were lysed by bead beating with the MP FastPrep system (MP Biomedicals) using lysing matrix B. DNA contamination of the RNA samples was removed by DNase I turbo (Invitrogen) digestion according to the manufacturer's protocol. The quantity and integrity of the DNase I-treated RNA was assessed using an Agilent 2100 BioAnalyzer according to the manufacturer's instructions (Agilent RNA 6000 Nano Kit). 1mg of crude isolated RNA was used for preparation of each small RNA libraries with TruSeq kit and detailed protocol provided by the manufacturer (Illumina). For total RNA-Seq, RNA samples were purified using AMPure XP magnetic beads (Becton Dickinson) with a 2.2:1 bead-to-RNA ratio. 2mgs of the purified RNA was used for total RNA library. The ribosomal RNA was removed with the riboPools Pan-Bacteria (siTools) according to the manufacturer's protocol. The Illumina-compatible RNA/cDNA sequencing libraries were prepared with the use of KAPA Stranded RNA-Seq kit (Roche) according to the detailed instructions provided by the manufacturer. For polyA selected libraries, the total RNA was incubated with poly(dT) beads, than washed extensively before recovering the enriched polyA fraction for further processing with VAHTS mRNA-seq V3 Library Prep Kit for Illumina® (Vazyme).